FIGURE 7.

Hsp31 and catalytic mutants prevent the formation of αSyn oligomers. A, flow chart depicting the fractionation of the αSyn fibrillization assay and detection of monomeric and oligomeric species. SDS-PAGE of the pellet and supernatant fractions was performed, and antibodies were used to detect αSyn followed by stripping of the membrane and reprobing with anti-HA tag antibody to detect Hsp31 (note that Hsp31 was purified using the MORF tag, but Protein A was removed proteolytically, and the HA tag is retained). B, Hsp31 suppresses the formation of SDS-resistant higher order oligomers detected with Western blotting of the pellet and supernatant fractions. C, Hsp31 catalytic triad mutants suppress αSyn fibrillization. All proteins were added at the same level of 2 μm. A.U., arbitrary units.